title: Eosinophil major basic protein activates human cord blood mast cells primed with fibroblast membranes by integrin-β1.
abstract: BACKGROUND
  Mast cell (MC) - eosinophil (Eos) activating cross-talk might be critical for the severity and chronicity of allergy. Among soluble mediators, eosinophil major basic protein (MBP), a hallmark of allergy, is particularly important because it was shown to activate specific MC subtypes. We previously demonstrated that MBP activates IgE-desensitized rat MC and human lung and cord blood-derived MC (CBMC) after priming with fibroblast membranal stem cell factor. However, a distinct mechanism for this activation was missing. Therefore, we aimed to investigate it.
  
  
  METHODS
  Major basic protein-1 activation of CBMC primed with fibroblast-derived membranes (FBM) was measured by β-hexosaminidase and tryptase release. Chemical cross-linking followed by micrometric flow cytometry probed direct interactions. Antibodies neutralized integrin-β1 and recognized its active form. Pertussis toxin (Ptx) was used to decrease integrin-β1 active form expression. Hematopoietic cell kinase (Hck) was identified by immunoprecipitation (IP) and silenced by siRNA.
  
  
  RESULTS
  Major basic protein-1-induced CBMC activation is mediated partly by MBP1-integrin-β1 interaction on the MC surface. FBM prime CBMC via a G protein, as confirmed by Ptx, to shift integrin-β1 to its active form. Following MBP1 binding, integrin-β1 binds Hck that further transduces the activation signal. MC priming with FBM leads to up-regulation in Hck protein level. MC integrin-β1 neutralization inhibits MBP1-induced activation and uptake. Hck silencing results with reduced MBP1-induced activation.
  
  
  CONCLUSIONS
  Fibroblast-derived membranes, integrin-β1, and Hck are involved in MBP1-induced activation of CBMC and therefore represent a distinct mechanism for this activation. This finding might implicate integrin-β1 and Hck as targets for decreasing MC - Eos activating cross-talk in allergy.
authors.affiliation: Department of Pharmacology & Experimental Therapeutics, Institute for Drug Research, School of Pharmacy, Faculty of Medicine, The Hebrew University of Jerusalem, Jerusalem, Israel.
authors.forename: M
authors.initials: M
authors.lastname: Ben-Zimra
authors.forename: I
authors.initials: I
authors.lastname: Bachelet
authors.forename: M
authors.initials: M
authors.lastname: Seaf
authors.forename: G J
authors.initials: GJ
authors.lastname: Gleich
authors.forename: F
authors.initials: F
authors.lastname: Levi-Schaffer
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000494:pharmacology; D057134:Antibodies, Neutralizing / Q000494:pharmacology; D002450:Cell Communication / Q000276:immunology; D002462:Cell Membrane / Q000276:immunology* / Q000378:metabolism; D047093:Eosinophil Major Basic Protein / Q000276:immunology* / Q000378:metabolism; D004804:Eosinophils / Q000276:immunology* / Q000378:metabolism*; D005347:Fibroblasts / Q000276:immunology* / Q000378:metabolism; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D020868:Gene Silencing; D006801:Humans; D019012:Integrin beta1 / Q000276:immunology* / Q000378:metabolism; D008407:Mast Cells / Q000276:immunology*; D051379:Mice; D011485:Protein Binding; D051572:Proto-Oncogene Proteins c-hck / Q000235:genetics / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D057134:Antibodies, Neutralizing; D019012:Integrin beta1; D051572:Proto-Oncogene Proteins c-hck; D047093:Eosinophil Major Basic Protein; D019206:GTP-Binding Protein alpha Subunits, Gi-Go
keywords: Hck; eosinophil major basic protein-1; fibroblast membrane fraction; integrin-β1; mast cells